1. Field of the Invention
The invention in the field of medicine and immunology, particularly innate immunity, relates to prevention and treatment of influenza infections with granulocyte-macrophage colony stimulating factor (GM-CSF) and to the prevention and treatment of acute pneumonia caused by other organisms.
2. Description of the Background Art
Seasonal influenza causes an estimated three to five million cases and 250,000 to 500,000 deaths worldwide annually. (See, for example, URL: who.int/mediacentre/factsheets/fs211/en/.) Pandemic disease can result in substantial additional morbidity, mortality and economic cost, as evidenced by the recent H1N1 swine influenza pandemic. The tremendous human burden of influenza mandates improved methods to prevent and treat this infection.
Control and clearance of influenza infection are believed to hinge on adaptive immunity, mediated by B and T lymphocytes. B cells produce antibodies to influenza hemagglutinin and neuraminidase, which protect against homologous virus (1). CD8+ cytolytic (or cytotoxic) T lymphocytes (CTL) cells clear influenza virus, limit viral replication and protect against lethal virus challenge (2-6). Recent studies also suggest a protective role for CD4+ T cells (7-10), which can lyse infected target cells (8), provide help to B cells, and promote expansion of CD8+ CTL (9). Based on the strength of the understanding that adaptive immunity is central to protection against influenza, preventive strategies have focused primarily on development of vaccines. Unfortunately, vaccines have been variably effective, in part because of antigenic shift and drift in the influenza viruses circulating in the population.
Recent studies have shown that innate immunity is also critical for resistance to influenza (11). Alveolar macrophages (AM) are the first line of host defense against respiratory microbes in general, and they contribute to clearance of influenza virus by Fc receptor-mediated phagocytosis (12). Depletion of AM markedly enhances disease severity caused by influenza in murine and porcine experimental models (13, 14). However, the mechanisms by which AM mediate protection are not well understood.
Granulocyte macrophage colony stimulating factor (GM-CSF) activity of was initially discovered in lung cell-conditioned medium where it stimulated growth of granulocytes and macrophages from cultured hematopoietic progenitors (Metcalf D. Blood 111:485-91, 2008). The nucleotide and amino acid sequences of murine and human GM-CSF (hGM-CSF) have been known for many years (Wong G G et al., Science 228:810-15 (1985); Lee, F, et al. Proc. Natl. Acad. Sci. USA 82:4360-4364, 1985; Miyatake, S et al., EMBO J 4:2561-68, 1985). hGM-CSF has been produced recombinantly in bacterial, yeast, mammalian, plant, and insect expression systems (see, fore example, Babu K S et al., Biotechnol Lett 31:659-64, 2009; Sardana R et al., Transgenic Res 16:713-21, 2008; Kim N S et al., Plant Mol Biol 68:263-75, 2008).
Recombinant hGM-CSF (rhGM-CSF) is most commonly used to promote hematopoietic recovery after cancer chemotherapy and bone marrow transplantation. This protein has additional biologic effects in activating immune responses to infection and inflammation, and in hematopoiesis (Sasaki M G et al., Vaccine 21:4545-9, 2003). rhGM-CSF has therefore been was used in clinical treatment of infectious disease, malignancies, wound healing and other conditions (Wang X L et al., Virus Res 143:24-32, 2009; Coon C et al., Scand J Immunol 70:106-15, 2009; Jin S et al., Cancer Biother Radiopharm 24:237-41, 2009; Lutzky J et al., J Immunother 32:79-85, 2009; Sato T et al., J Clin Oncol 26:5436-42, 2008; Dai S et al., Mol Ther 16:782-90, 2008; Mann A et al., J Investig Dermatol, Symp Proc 11:87-92, 2006)
Therapeutic antiviral activity has been observed with rhGM-CSF used as an immunological adjuvant or in combination with antivirals (Sasaki et al., supra; Elias E G et al., Cancer Biother Radiopharm 23:285-91, 2008; Qiu J T et al., Vaccine 25:253-63, 2007; Zhai Y Z et al., Intervirology 52:152-63, 2009). Antiviral effects of rhGM-CSF alone against hepatitis B virus (Martin J et al. Hepatology 1993; 18:775-80, 1993), HIV (Matsuda S et al., AIDS Res Hum Retrovir 11:1031-8, 1995) and Herpes simplex virus (Altamura M, et al., Immunopharmacol Immunotoxicol 19:425-36, 1997) have been reported.
GM-CSF contributes to maturation of mononuclear phagocytes and AM (15, 16). In patients with pulmonary alveolar proteinosis, circulating neutralizing antibodies against GM-CSF cause AM dysfunction (17), and AM from GM-CSF-deficient (GM−/−) mice have impaired capacity for phagocytosis and cytokine production, which functions were restored by GM-CSF treatment (18). Studies in GM−/− mice showed that GM-CSF contributed to immune responses during pneumonia that was caused by Pseudomonas aeruginosa and Pneumocystis carinii; administration of GM-CSF to septic patients reversed monocyte immunosuppression and improved the clinical course (19-21). DNA vaccination with a plasmid encoding influenza hemagglutinin, GM-CSF and IL-12 resulted in reduced viral titers and increased neutralizing antibody titers (22), suggesting that GM-CSF may enhance adaptive immune responses.
Huang H et al., showed protective effects of recombinant human GM-CSF on H1N1 influenza virus-induced pneumonia in mice (23).
Secondary bacterial infection occurs commonly after pulmonary virus infection and can cause severe disease in humans. Mechanisms responsible for this “synergy” in the lung are poorly understood. K. Sun and D W Metzger reported (Nature Med. 14:558-564 (2008)) that in mice, pulmonary interferon-γ (IFNγ) produced during the T cell response to influenza infection inhibited initial bacterial clearance from the lung by AM. This suppression of phagocytosis which correlated with lung IFNγ levels but not with viral burden, resulted in enhanced susceptibility to secondary pneumococcal infection. This effect could be prevented by neutralizing IFNγ after influenza infection. Thus, while promoting induction of anti-influenza adaptive immunity, this T cell product of the immune response, IFNγ, suppressed innate protection against extracellular bacterial pathogens in the lung.
In response to the need in the art for improved approaches to prevent and treat influenza infections, the present inventor conceived of the present invention.
(Certain references above and below appear as parenthetical numbers and appear in a reference list. Others are provided directly in the body of the text.) Citation of the above documents is not intended as an admission that any of the foregoing is pertinent prior art. All statements as to the date or representation as to the contents of these documents is based on the information available to the applicant and does not constitute any admission as to the correctness of the dates or contents of these documents.